Viral infection is implicated as a possible risk-factor for neurodegenerative diseases, including Parkinson's disease (PD). Infection with neurotropic alphaviruses can produce many of the same long-term degenerative effects seen in PD, including protein aggregation, increased levels of oxidative stress, autophagy/mitophagy defects, neuroinflammation, and neuronal death. Neurotropic viruses may therefore represent a better animal model for studying gene x environment interactions in PD that recapitulate more features of the human disease than drug-based lesioning models or genetic models that often lack a pronounced phenotype in the substantial nigra. Evidence suggests that viral infection may act in synergy with other recognized risk-factors, such as aging, genetic factors, and/or previous exposures to environmental neurotoxins to promote neurodegeneration. Among neurotoxins that could have a pronounced gene x environment interaction with neurotropic viruses, Manganese (Mn) is of interest because excessive exposure early in life can have lasting effects on neurological function and can also enhance the neurovirulence of alphaviruses. Here, we propose a novel method for testing gene x environment interactions in Parkinsonism using a neurotropic alphavirus expression system (AES). Following convenient intranasal inoculation, 100% of animals become infected and the progression of infection can be non-invasively monitored in situ using luciferase-expressing viruses and whole body bioluminescence imaging. Neuroinvasion occurs through olfactory sensory neurons and the infection spreads along the neuronal axis in a pattern that mimics the Braak-staging system of PD. The severity and persistence of viral infection can be tightly controlled and our preliminary data using unbiased stereology indicate that AES infection results in significant loss of dopaminergic neurons in the substantial nigra pars compacta (SNpc). It is the Central Hypothesis of this proposal that intranasal infection with luciferase- expressing alphavirus will result in neuroinvasion through olfactory and subcortical structures resulting in progressive loss of dopaminergic neurons in the substantia nigra that is associated with inflammatory activation of glial cells. Moreover, we postulate that pre-exposure to Mn during juvenile development will enhance susceptibility to viral-mediated loss of dopaminergic neurons, in part through an increased neuroinflammatory response. This hypothesis will be tested in two Specific Aims during the 2-year project period as follows: Specific Aim 1 - Determine the optimal viral titer and phenotype for inducing persistent infection and neurodegeneration in the basal ganglia following intranasal installation of luciferase-expressing Western Equine Encephalitis Virus (WEEV); Specific Aim 2 - Characterize the disease phenotype resulting from exposure to Mn pre- and post-viral infection. We expect that these studies will demonstrate that the proposed AES-based model is a powerful method for inducing parkinsonism in mice, as well as for transgene delivery into the CNS, that can be used to identify novel gene x environment interactions relevant to PD. Additionally, we expect that exposure to Mn during juvenile development will exacerbate neurodegeneration following adult infection with WEEV. The results of these studies will increase our understanding of the environmental links to neurodegenerative disease and will provide a powerful new animal model for studying virus/toxin interactions in the CNS.